APPLICANT'S DESCRIPTION: Cajal bodies (coiled bodies, CBs) are nuclear organelles whose structural morphology and molecular composition have been conserved from plants to animals. Although their precise functions have yet to be determined, CBs are involved in the biogenesis of small nuclear ribonucleoproteins (snRNPs). CBs contain not only high concentrations of partially mature snRNPs, but components of the basal transcription and cell cycle control machineries as well. Intriguingly, CBs associate with specific chromosomal loci in interphase human cells, including various snRNA genes and the two cell cycle-dependent histone gene clusters. The major thrust of this proposal is to understand the mechanics and dynamics of the interactions between genes and CBs at both the cellular and molecular levels. Previous analysis has shown that CBs colocalize with clustered and single-copy snRNA genes and that the association appears to be mediated by nascent snRNA transcripts. A series of experiments is proposed to ascertain additional functional requirements for these interactions through the use of stably transfected arrays of snRNA and histone gene constructs. Specific Aims of this proposal are: (1) to identify functional elements within CBs and nascent U2 snRNAs that are required for their mutual association, (2) to understand the dynamics of CB formation and snRNA gene interaction in living cells using a novel BAC-based vector approach to tag an artificial array of U2 genes in conjunction with the GFP-lac repressor/operator system and (3) to identify chromosomal elements within histone gene clusters responsible for both interaction with CBs and for recruitment of a histone gene transcriptional activator.